Quantifying actin dynamics in developing Drosophila neurons using LoG filters

Authors

• KM O'Neill
• LJ Campanello
• J Corbett-Frank
• AY Clarke
• HY Fang
• W Losert
• E Giniger

Abstract

The phenomenon of growth cone motility and the associated contributions of cytoskeletal proteins have been studied extensively in vitro, where growth cones are often characterized as having palm-like (lamellipodial) structures. However, the Giniger laboratory has discovered a novel form of growth cone motility in vivo in the TSM1 axon of Drosophila, in which the growth cone is more filamentous (filopodial) in structure and appears to be driven by dynamic rearrangement of actin. In this work, we aim to elucidate how dynamic actin rearrangement contributes to the advancement and morphology of this filopodial growth cone in vivo. We have developed an image processing method that employs 3D Laplacian of Gaussian (LoG) filters to automatically segment the TSM1 growth cone from surrounding tissue in images obtained by fluorescent live imaging. In brief, this is accomplished by applying the LoG filter of various sizes and rotations to the 3D image. We then use the medial axis skeleton of the processed growth cone image to quantify the distribution of actin along the main body of the axon. Currently, we are extending the analysis to include tracking of minor protrusions over time, a task that is laborious when performed by hand. This image analysis method represents a general approach for successfully segmenting filamentous, 3D shapes from noisy backgrounds and has broad applicability to other fields. The ultimate goal of perfecting this method is to quantify the actin dynamics that underlie growth cone morphology and advancement of TSM1 in vivo.

Scientific Focus Area: Cell Biology

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